Table 3 Application of some interesting bacterial laccases that degrade different compounds and may be useful in water treatement

Pharmaceutical compounds

 Recombinant laccase from Yersinia enterocolitica expressed in E. coli

CS

Expressed in E. coli with IPTG induction

Degradation of non-steroidal anti-inflammatory drugs

E. coli cells harboring laccase from Y. enterocolitica were treated with 0.1 mM Tween 80 and CuCl₂ 0.2 mM in buffer pH 6, at 45 °C for 30 min. After that, diclofenac and aspirin were added at 5 mg/L incubated at 45 °C

After 24 h, both diclofenac and aspirin were fully degraded

In the case of the diclofenac, laccase oxidation by hydroxylations of 4′ or 5′ positions of the second benzene ring could be the modifications

[136]

 Streptomyces cyaneus

C

Production of laccase was done in ISP9 mineral medium, with soy flour (10 g/L) as carbon source and a copper concentration of 1 mg/L CuSO₄∙5 H₂O. Cultures were incubated at 30 °C for 23 days. Cell-free culture supernatant was collected, filtered and stored as enzyme source

Degradation of non-steroidal anti-inflammatory drugs (diclofenac: DFC) and mefenamic acid: MFA)

The reactions were performed in citrate phosphate buffer (30–40 mM) at three different pH values (5, 6 and 7), with a pollutant concentration of 20 mg/L. 2000 U/L of crude enzyme preparation were used. The reactions were incubated in the dark at 25 °C for 12 days

The enzyme showed a high conversion rate under acidic conditions (pH 5, 6), with 50% of conversion after 2 days for DFC. With respect to MFA, the highest conversion was obtained in pH 6

Not discussed

[137]

 Recombinant Streptomyces ipomoea SilA laccase expressed in E. coli

F

2 L of LB medium at 37 °C were inoculated with 40 mL of an exponential-growth-phase culture. When exponential growth had resumed, the temperature was reduced to 28 °C, and SilA expression was induced with 1 mM IPTG. Purified enzyme was used

Degradation of fluoroquinolone antibiotics (ciprofloxacin: CIP, and norfloxacin)

The reactions were carried out in 50 mM phosphate buffer pH 8 at 35 °C, using 0.4 U/mL of laccase and 50 μg/mL of each fluoroquinolone. Several mediators at concentrations of 0.1, 0.3 and 0.5 mM were tested

After 24 h and with 0.5 mM acetosyringone, higher than 90% percent conversions were obtained for both antibiotics, with a detoxification effectiveness of 70% for CIP and 90% for norfloxacin

Possible oxidation of piperazine substituents

[140]

 Streptomyces mutabilis A17

F

The culture was done in a solid-state fermentation, using cotton seed cake (5 g/L) as substrate, supplemented with mineral salts and glucose 1% (w/v). The medium was inoculated with a spore suspension and incubated for 6 days at 35 °C. The laccase was extracted and purified

Degradation of sulfa antibiotics (sulfadiazine and sulfathiazole)

In a 100 mM citrate–phosphate buffer pH 6 were dissolved each sulfa drug, with a final concentration of 50 mg/L. To this solution were added the laccase (81.3 U/mg), and 1 mM HBT (mediator). The reaction was done at 50 °C for 60 min

Under the conditions previously described, 73 and 90% removal efficiencies were achieved to sulfadiazine and sulfamethoxazole solutions, respectively. Moreover, the reaction products showed less antibiotic effect in bacterial cultures

Not reported

[138]

Dye based pollutants

 Recombinant laccase from Klebsiella pneumoniae expressed in E. coli

F

E. coli cells were grown in LB medium at 37 °C until 0.6–0.8 DO. After that, were induced with IPTG for 20 h at 16 °C. Purified laccase

Decolorization of synthetic dyes

The reactions were done with 0.025, 0.05 and 0.1 U of the purified enzyme, in 50 mM citrate–phosphate buffer (pH 4.0 and 7.5) and 15 μL dye solution (100 mg/L) at 70 °C in 90 min

All the 10 dyes tested were efficiently oxidized under by the enzyme alone in both acidic and neutral conditions

Not reported

[99]

 Recombinant E. coli K-12 CueO expressed in Pichia pastoris

F

48 h culture in BMGY at 28 °C. The induction was made with methanol 1% and 0.2 mM CuSO₄ for 144 h, feeding methanol each 24 h. Purified laccase

Decolorization of synthetic dyes

The reactions were carried out at 55 °C in phosphates buffer 50 mM, pH 7.5, with a dye concentration of 80 mg/L and supplemented with 1 mM CuSO₄ and 0.1 mM of acetosyringone as mediator. 1 μL of purified laccase was used

After 3 h of reaction, the laccase decolorized almost all the Congo red and malachite green tested. After 24 h, 90% of the remazol brilliant blue R were degraded

Not mentioned

[128]

 Recombinant and mutant laccase WLF from Bacillus pumilus expressed in E. coli

F

Culture grown at 37 °C in LB medium until 0.5 DO. After that, were added IPTG (0.4 mM) and CuSO₄ (0.25 mM) and maintained at 15 °C for 24 h. Purified laccase

Decolorization of synthetic dyes

Reaction mixture consisted in 0.25 mg of dye, 2 mg/L of purified laccase 1 mM de acetosyringone in 5 mL of 100 mM carbonate buffer pH 10, at 37 °C

Highest transformations of all the dyes tested. The efficiency with aromatic heterocyclic dyes was lower compared with azo, anthraquinonic and triphenylmethane dyes

Not reported

[125]

 Bacillus safensis S31

SS

B. saensis cells were cultured on nutrient agar sporulation medium and incubated at 35 °C for 4 days. After that incubation time, spore suspension was prepared, used as a source of laccase

Decolorization of synthetic dyes (malachite green, toluidine blue and reactive black 5)

To 2 mL of 50 mM acetate buffer (for pH values of 3–6) or 50 mM Tris buffer (for pH values of 7 and 8) were added the spore laccase suspension (8 U/L) and dye (final concentration of 10 mg/L). The effect of ABTS (15 μM) as mediator was also studied. The reactions were carried out at 30 °C for 2 h

Almost all the oxidation conditions showed better results with ABTS. The highest decolorization values for malachite green and toluidine blue were achieved between 5 and 7 pH values, while with reactive black were between pH 3 and 5

Not mentioned

[134]

 Recombinant Thermus thermophilus SG0.5JP17-16 expressed in Pichia pastoris

F

An inoculum of Pichia pastoris cells growth in BMGY medium were used to inoculate BMMY medium containing 0.1 mM CuSO₄. The culture was cultivated at 30 °C for 7 days with daily addition of 1% methanol. The enzyme was purified from the supernatant

Decolorization of synthetic dyes (reactive black B, reactive black WNN, congo red and remazol brilliant blue R)

A reaction mixture of 50 mM phosphates buffer pH 7.5, 10 μM CuSO₄, 50 mg/L dye and 40 U/L of purified laccase were heated at 70 °C for 24 h

After 24 h the decolorization efficiency for congo red, reactive black B and reactive black WNN was higher than 90%, while for remazol brilliant blue R was around 70%

Not mentioned

[126]

 Recombinant Streptomyces ipomoeae SilA, expressed in E. coli

F

E. coli BL21 (DE3) transformed and containing the codifying gene of SilA

2 L of Luria–Bertani (LB)

Decolorization of synthetic dyes

Laccase SilA and three mediators (0.1 mM), acetosyringone (AS), syringaldehyde (SA) and methyl syringate (MeS), by 24 h at 35 °C pH 8, and different dyes (acid black 48: AB48, acid orange 63: AO63, reactive black 5: RB5, orange II: OII, tartrazine: TART, azure B: AB, indigo carmine: IC, cresol red: CR

Laccase and mediators such as AS and MeS enhanced the decolorization and detoxification of a variety of textile dyes, principally RB5, OII, and IC, diminishing the toxicity of acid orange 63, tartrazine

The oxidation of MeS (which has the weakest acceptor group at the para-position) gives an stable phenoxy radical

[139]

Plastic and polycyclic aromatic hydrocarbons (PAHs) compounds

 Recombinant B. subtilis CotA expressed in E. coli

F

E. coli cells harboring the plasmid with the CotA gen were grown at 37 °C in LB medium. When the culture reaches 0.6 DO, were added IPTG and CuSO₄ to final concentrations of 0.1 and 0.25 mM, respectively. The incubation temperature was reduced to 25 °C for 6 h. After that, the culture agitation was stopped for 12 h. Purified laccase was used

Degradation of PAHs (anthracene, pyrene benzo[α]pyrene, phenanthrene, fluoranthene, etc.)

The reactions were carried out in 50 mM acetate buffer pH 4 with 10% acetonitrile, with PAHs concentrations from 0.1 to 1 mg/L and laccase concentration of 3 U/mL. The reactions were incubated for 24 h at 20, 40 and 60 °C

Just anthracene and benzo[α]pyrene were significantly oxidized (almost complete oxidations at 60 °C), the other ones had degradation values from 0 to 40% in all the conditions tested

Not reported

[124]

 Streptomyces cyaneus

C

Production of laccase was done in ISP9 mineral medium, with soy flour (10 g/L) as carbon source and a copper concentration of 1 mg/L CuSO₄∙5 H₂O. Cultures were incubated at 30 °C for 23 days. Cell-free culture supernatant was collected, filtered and stored as enzyme source

Degradation of bisphenol A

The reactions were performed in citrate phosphate buffer (30–40 mM) at three different pH values (5, 6 and 7), with a pollutant concentration of 20 mg/L. 2000 U/L of crude enzyme preparation were used. The reactions were incubated in the dark at 25 °C for 12 days

Under all the conditions tested after 2 days there was full degradation, especially at pH 5 and 6

Not reported

[137]